This research proposal will determine whether, a distinct subpopulation of frontonasal mesenchyme identified by GAP-43 immunoreactivity in the mouse, is responsible for producing RA-inductive signals that regulate olfactory system morphogenesis. In PAX-6 mutant mice the olfactory system fails to develop, and this sub-population of GAP-43 immunoreactive mesenchymal cells is absent from the mutant PAX-6 mouse's frontonasal mesenchyme. Accordingly this study will test the hypothesis that, the GAP-43 immunoreactive mesenchymal cells are a distinct source of retinoid inductive signals that lead to the morphogenesis of the olfactory system. This hypothesis will be tested by addressing the following four specific aims. Specific Aim 1. To determine the position of the GAP-43 immunoreactive mesenchymal cells in the normal mouse. These experiments will show whether the GAP-43 immunoreactive mesenchymal population is located sufficiently close to retinoid responsive forebrain regions to provide the retinoid inductive signals. Specific Aim 2. To determine whether the GAP-43 immunoreactive mesenchymal cells produce retinoid signals. These studies will identify the source or sources of retinoid signals in the frontonasal mesenchyme and determine their relationship to the GAP-43 immunoreactive population of cells. Specific Aim 3. To assess the origin of the GAP-43 immunoreactive mesenchymal cells of the frontonasal mesenchyme. It is likely that the GAP-43 immunoreactive cells in the mouse frontonasal mesenchyme are derived from the neural crest since during development cranial crest cells migrate tot he frontonasal region (Matsuo et all., 1993). Therefore, populations of frontonasal mesenchyme that are neural crest derived will be identified by immunostaining the frontonasal mesenchyme with antibodies that recognize neural crest specific antigens. Specific Aim 4. To assess potentially ectopic sites of retinoic acid production along the migratory pathway of th neural crest to the frontonasal region in mutant PAX-6 mice. Since PAX-6 influences migration of neural crest to the frontonasal mesenchyme (Matsuo et al., 1993), these studies will determine whether the population of GAP-43 immunoreactive mesenchymal cells is ectopically positioned due to aberrant neural crest migration in PAX-6 mutant mice. In the embryonic mouse, the frontonasal mesenchyme is made up of a mosaic of cells, including a large population of post-migratory cranial neural crest cells (Le Douarin, 1983; 1986; 1993; Noden, 1975; 1988; Richman and Tickle, 1989; Serbedzija et al., 1992). Therefore, distinct subpopulations of ventrolateral mesenchyme may be derived from the neural crest. Specific Aim 1 will define the location of these mesenchymal populations relative to the developing olfactory system. Specific Aim 2 will determine whether the GAP-43 immunoreactive mesenchymal cells or any of the other antigenically distinct subpopulations of frontonasal mesenchyme provide RA signals during olfactory system development. Specific Aim 3 will then further characterize this retinoid-producing population of mesenchyme, by examining whether these cells share antigenic properties of neural crest cells indicative of their lineage. Since mutant PAX-6 animals show aberrant migration of neural crest as well as failed development of their olfactory systems, if the migrating neural crest cells are the source of RA-signals, then sites of ectopic RA production would be expected in the mutant PAX-6 mice. Accordingly mutant PAX-6 mice are an excellent system in which to test this hypothesis. Thus this work may define a distinct and transient sub lineage of neural crest cells distinguished by GAP-43 immunoreactivity that populates the frontonasal mesenchyme and is specialized to produce region specific inductive signals that regulate vertebrate forebrain morphogenesis.